These studies represent a detailed investigation of the regulation of hepatic fatty acid synthesis and metabolism by insulin and other hormones. We and others have presented evidence that the hormonal regulation of hepatic acetyl-CoA carboxylase may underly the acute modulation of both hepatic acid synthesis and metabolism. These studies are designed to describe the major mechanisms by which insulin and glucagon regulate enzyme activity. We plan to investigate in isolated hepatocytes the influence of insulin and glucagon on the activity of acetyl-CoA carboxylase (ACC) simultaneous with effects on the phosphorylation state of the enzyme, on the intracellular concentration of allosteric effectors and on the protomer-polymer equilibrium of the enzyme. The phosphorylation sites of the enzyme as phosphorylated in the whole cell and changes in the occupancy of these sites in response to insulin and glucagon will be described. The major protein kinases and phosphatases that mediate changes in the phosphorylation state will be characterized. We will also apply laser light scattering spectrometry to investigate the relationship between alterations in covalent phosphorylation and pertubations of the protomer-polymer equilibrium of the enzyme in real time. We will the influence of insulin and glucagon on the activity of ACC protein kinases and phosphatases. Future studies will attempt to determine whether protein-free hepatocyte cytosol contains "messages" that influence the activity of either kinase or phosphatase. These descriptions will not only provide a more detailed description of the hormonal regulation of hepatic fatty acid metabolism, but may provide a model system for the study of the molecular mechanism of insulin action. Such information is needed to understand the misregulation of fatty acid metabolism in diabetes mellitus.